The complete chloroplast genome of Ligusticopsis acaulis (Shan et Sheh) Pimenov (Apiaceae), an endemic species from China

Abstract Ligusticopsis acaulis, belonging to the family Apiaceae (Umbelliferae), is endemic to China. The complete chloroplast genome sequence of L. acaulis was assembled and annotated for the first time in this study. The results showed that the plastome was 148,509 bp in length and consisted of a pair of inverted repeat regions (IRs: 19,468 bp), a large single-copy region (LSC: 91,902 bp), and a small single-copy region (SSC: 17,671 bp). A total of 114 unique genes were annotated, including 80 protein-coding, 30 tRNA, and four rRNA genes. According to the phylogenetic analysis, L. acaulis belongs to the tribe Selineae, with a close relationship to Ligusticum hispidum (Franch.) Wolff.


Introduction
Ligusticopsis was originally established by Leute in 1969, with the genus containing 14 species from China (Leute 1969). Ligusticopsis acaulis, a species endemic to China, has diverse chemical composition and is used as a substitute for Peucedani Radix in the Yunnan Province (Rao et al. 1995). The species was first described by Shan et al. (1986) and was named Peucedanum acaule Shan et Sheh. Pimenov (2017) classified Ligusticopsis acaulis as a synonym of P. acaule. The genus Peucedanum L. (Apiaceae) contains approximately 120 species widely distributed in Eurasia and Africa (Pimenov and Leonov 1993). There are about 40 species of Peucedanum in China, with 33 species being endemic (Sheh and Watson 2005). However, due to the complex species composition and ambiguous relationship with related taxa, the taxonomy of Peucedanum has long been contentious.
In recent years, the chloroplast (cp) genome was applied to the phylogenetic reconstruction of Apiaceae, whereby a few contentious relationships were successfully resolved. Here, the complete cp genome sequence of L. acaulis was characterized, and its phylogenetic relationships with related taxa were established.

Materials and methods
Sample collection, DNA extraction, and sequencing The specimen was collected from the Sea Grass Mountain in Huize, Yunnan Province of China (26 23 0 99 00 N, 103 24 0 01 00 E, and altitude 3570 m), and the voucher was deposited in the School of Pharmaceutical Sciences and Yunnan Key Laboratory of Pharmacology for Natural Products, Kunming Medical University (appraiser: Jing Zhou; contact person: Jing Zhou, zhoujing_apiaceae@163.com) under voucher number LZ0901 ( Figure 1). The modified CTAB method was used to extract total genomic DNA from leaf tissue (Doyle and Doyle 1987).

Chloroplast genome assembly and annotation
The genome was sequenced using the Illumina HiSeq 2500 platform (Majorbio, Shanghai, China) with the paired-end (2 Â 300 bp) library. The raw reads were filtered using Trimmomatic with default parameters set to remove the adapter and low-quality sequences (Bolger et al. 2014). Clean data were then assembled into circular contigs using GetOrganelle v.1.7.5.3 (Jin et al. 2020). Finally, using Peucedanum praeruptorum Dunn. (GenBank accession number: MN016968) as a reference, the plastome was annotated by PGA (Qu et al. 2019) and manually modified in Geneious (Kearse et al. 2012). The circular genome map was drawn by CPGView (www.1kmpg.cn/cpgview/).

Phylogenetic analyses
A total of 31 published plastome sequences of Apiaceae from NCBI were acquired to investigate the phylogenetic position of L. acaulis. All sequences were aligned using MAFFT (Katoh and Standley 2013) with Bupleurum boissieuanum H. Wolff as the outgroup. The molecular phylogenetic tree was constructed using the maximum-likelihood (ML) analysis in RAxML (Stamatakis 2014) with 1000 rapid bootstrap replicates.

Phylogenetic analysis
To confirm the phylogenetic position of L. acaulis, an ML phylogenetic tree was constructed using 31 cp genome sequences from NCBI. Using MAFFT, sequences were aligned. B. boissieuanum served as the outgroup. Our phylogeny showed that L. acaulis was closely related to Ligusticum hispidum and belonged to the tribe Selineae ( Figure 3). Meanwhile, the phylogenetic inference provided an opportunity to delimit the species of the genus Ligusticopsis and understand their evolutionary relationships. The complete cp genome of L. acaulis provided important information for the phylogenetic analysis of the genus Ligusticopsis and laid the foundation for further reconstruction of the Apiaceae phylogeny.

Author contributions
Zhenwen Liu and Jing Zhou conceived and designed the study. Junmei Niu, Xinyue Wang, Jiarui Yue, and Shilin Zhou analyzed the experimental results and interpreted the data. Junmei Niu drafted the manuscript. Zhenwen Liu and Jing Zhou revised the manuscript. All authors have read and approved the final manuscript.

Ethical approval
Investigations of the authors with human participants or animals are not included in this article. Hence, no ethical approval or permissions were required.